The Investigation Of Interreactions Between The Virb4 Protein Of Brucella And The Bovine Embryo Trophoblastic Cells

Brucellosis is a difficult zoonotic disease in many parts of the world, although we have known it more than before. The harm of it is not optimistic with over half a million new cases each year. These pathogens can cause abortion and sterility inanimals, while in humans the disease is characterized by recurrent episodes of fever. The TFSS is relationship to the brucella's infest, survival in the cell and multiplication, it is the important virulence factor. It is significant to understand the conjunctive mechanism between the TFSS and the rephocyte's target proteinum.First, To investigate epidemic of brucellosis in Xinjiang Province,1859 samples of livestock were detected by agglutination test from 27 different territory including 1425 of sheep serum and 434 cattle serum, The date indicated that the infection was fairly critical in infected territory. The maxlmum masculine rate was 31.51%. The average masculine rate was 8.42% in sheep and was 6.68% in cattle.This state is same to the world.Then the target proteins of the bovine embryo trophoblastic cells which conjunctive with virB4 gene were screened through yeast two-hybrid. And detect the mRNA of the proteins by Real Time Fluorescence Quantitative PCR. To amplify virB4 from RB51 vaccine by PCR species-specific primers were designed of the virB4 gene. Through ligate the insert with the cloning vector, transformation, DNA sequencing. After that digested by Nde I/Pst I, ligate it with the expression vector DNA sequencing, restriction enzyme analysis, transformed pGBKT7-virB4 into Y187. After that, test the transcription activation and toxicity. The cDNA library of rephocyte infected with vaccine RB51 strain was constructed, Then screened the target protein which interacted with virB4 gene by yeast two-hybrid. Analysising them by Blastn to determined the factions. At last, detect the mRNA of the protein by Real Time Fluorescence Quantitative PCR. Result:the genes virB4 contained the entire genes coding region. Recombinant plasmid pGBKT7-virB4 was cloned into the Y187 successfully.13 kinds of proteins were screened. Choosed two proteins Q10 and SLC3A2 to detect by Real Time Fluorescence Quantitative PCR, both of their expression were increased. Increasing of Q10 will promote the cell's metabolism. Increasing of SLC3A2 will change the transport of amino acids. In this study, it is useful to give a upstream work for researching the molecular mechanism of abortion.